Redirecting the substrate specificity of heparan sulfate 2-O-sulfotransferase by structurally guided mutagenesis
Redirecting the substrate specificity of heparan sulfate 2-O-sulfotransferase by structurally guided mutagenesis
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DOI：
10.1073/pnas.0806975105
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发表时间：
2008-12-02
影响因子：
11.1
通讯作者：
Pedersen, Lars C.
中科院分区：
文献类型：
--
作者：
Bethea, Heather N.;Xu, Ding;Pedersen, Lars C.
硫酸乙酰肝素(HS)是一种参与调节细胞生长和凝血等重要生理功能的多糖。HS的生物合成涉及多种特殊的磺基转移酶，如2-O-磺基转移酶(2OST)，它将磺基转移到HS中的艾杜糖醛酸(IDOA)或葡萄糖醛酸(GLCA)的2-羟基位置上。在这里，我们报道了鸡2OST与3‘-磷酸腺苷5’-磷酸形成的络合物的高三聚体晶体结构。基于结构的突变分析已经确定了与底物特异性有关的氨基酸残基。突变体R189A只将硫酸盐转移到多糖中的GLCA部分，而突变体Y94A和H106A优先将硫酸盐转移到IOA单位。我们的结果证明了操纵2OST的底物专一性来合成具有独特硫化模式的HS的可行性。这项工作将有助于开发一种酶促合成肝素类药物的方法。
Heparan sulfate (HS) is a polysaccharide involved in essential physiological functions from regulating cell growth to blood coagulation. HS biosynthesis involves multiple specialized sulfotransferases such as 2-O-sulfotransferase (2OST) that transfers the sulfo group to the 2-OH position of iduronic acid (IdoA) or glucuronic acid (GlcA) within HS. Here, we report the homotrimeric crystal structure of 2OST from chicken, in complex with 3'-phosphoadenosine 5'-phosphate. Structural based mutational analysis has identified amino acid residues that are responsible for substrate specificity. The mutant R189A only transferred sulfates to GlcA moieties within the polysaccharide whereas mutants Y94A and H106A preferentially transferred sulfates to IdoA units. Our results demonstrate the feasibility for manipulating the substrate specificity of 2OST to synthesize HS with unique sulfation patterns. This work will aid the development of an enzymatic approach to synthesize heparin-based therapeutics.